The overall goal of this project is to understand how the shape of cells in the context of tissue organization controls transmembrane signal transduction. In the previous term we had found that cells with different aspect ratios produce transient microdomains of activated receptors and differentially regulate cytoplasmic and nuclear signaling. Building on these observations in the coming term we plan to use a combination of constrained analytic representations and numerical simulations with super resolution imaging of biochemical reactions in individual cells assembled within engineered 3D chips to mimic tissue organization to obtain deep understanding of transmembrane signaling when the cell is part of the 3D tissue environment. Our central hypothesis is that within tissues, cells, to maintain their functional phenotype sense a multifaceted relationship between extra and intra cellular spaces and density of receptors at the cell surface, in the context of various cell shapes. This multidimensional relationship controls transmembrane signal transduction and cell health. To test our central hypothesis we propose to develop analytical models of role of cell shape in information storage and retrieval as cells function within tissues. We will address one key question: 1) how changes in intra and extracellular spaces and consequently their effect on concentrations of soluble components such as receptor agonists, and intracellular adapters and transducers separated by the plasma membrane and membrane bound receptors interact to regulate transmembrane signaling. Using human and rat vascular smooth muscle cells as model systems we will use the analytical representations to develop multicompartment ODE and PDE models and run numerical simulations to determine how relationships between intracellular and extracellular spaces and cell shape control transmembrane signal transduction. To test model predictions we will develop 3D chips ? microfabricated devices that enable the assembly of multilayered cells of different shapes and varying extracellular spaces to mimic organization of these cells within tissues. .We will then test model predictions of control of formation of activated receptor signaling complexes using super resolution fluorescence microscopy and single molecule fluorescence spectroscopy. From these studies we hope to elucidate fundamental principles of how cell shape and tissue organization information is processed at the transmembrane level.